This was a double-blinded, randomized, placebo-controlled evaluation of the prophylactic antimalarial activity of atovaquone (a broad spectrum antiprotozoal agent) in sixteen healthy volunteers with induced Plasmodium falciparum malaria. The study was conducted on an outpatient basis, and had three arms: a) six volunteers receiving 750 mg qd starting the day before challenge; b) six volunteers receiving a single dose of 250 mg the day before challenge; and c) four volunteers receiving placebo. Volunteers were challenged with the NF54 strain of chloroquinesensitive P. falciparum, by the bites of infected Anopheles stephensi mosquitoes. Efficacy was evaluated by serial blood examinations (on 35 occasions over a three month period) for malaria parasites and by monitoring for symptoms and signs of infection. Parasites were cultured and their sensitivity to atovaquone was determined. All four placebo recipients developed parasitemia and were successfully treated with chloroquine. All twelve drug recipients remained free of circulating parasites. Thus, both high and low dose atovaquone provided effective prophylaxis [95% Cl 0.61-1.00; p = 0.005]. However, in the low dose recipients, plasma levels by 6.5 days after challenge (the earliest anticipated appearance of P. falciparum in blood) had fallen well below concentrations that failed against erythrocytic parasites in previous studies. This indicates the organisms were killed prior to day 6.5, while still in the liver. There was no evidence for a partial therapeutic response in the form of subclinical parasitemia or a prolonged hepatic phase. We conclude that Atovaquone protects non-immune subjects against mosquito-transmitted falciparum malaria. Furthermore, the evidence indicates there is a causal component against liver stage parasites. This extra dimension of antiparasitic activity eliminates the requirement for weeks of post- exposure therapy currently necessary with chloroquine or mefloquine, which have suppressive, but not causal, antimalarial effects. Atovaquone, as a component of Malarone, thus offers a much-needed new therapeutic approach to malaria chemoprophylaxis.